The endosomal-lysosomal pathway is a major proteolytic system in neurons as well as in other eukaryotic cells. The importance of this pathway in neuronal signaling and synaptic function is highlighted by recent findings that disturbed endosome-to-lysosome trafficking in Drosophila mutants results in abnormal synaptic growth and impaired neurotransmission. Furthermore, aberrations in this pathway have been implicated in the pathogenesis of a number of neurological disorders and neurodegenerative diseases, including Alzheimer's disease, Huntington's disease, and more than 40 lysosomal storage disorders. Despite the critical importance of the endosomal-lysosomal pathway in normal physiology and diseases, the molecular mechanism that controls endosome-to-lysosome trafficking remains poorly characterized. The long-term goal of this research is to understand, at the molecular level, how endocytosed proteins are sorted and transported to lysosomes for degradation, and how this process becomes dysregulated in neurological and neurodegenerative diseases. Recent work by the applicant and others has revealed a crucial role for the endosomal protein hepatocyte growth factor-regulated tyrosine kinase substrate (Hrs) in regulating endosome-to-lysosome trafficking. However, the mechanism of action of Hrs remains unclear. The applicant's preliminary studies have identified three Hrs-interacting proteins, sorting nexin 1 (SNXl), signal transducing adaptor molecule (STAM), and huntingtin-associated protein 1 (HAP1). This project will test the hypothesis that Hrs and its associated proteins SNXl, STAM, and HAP1 are key components of the endosomal trafficking machinery that control the sorting and trafficking of endocytosed proteins to lysosomes for degradation. A combination of biochemical, proteomic, molecular biological, and cell biological approaches will be used to characterize Hrs-associated protein complexes and determine their roles in ubiquitin-dependent endosome-to-lysosome trafficking and in neurodegeneration. Results from these studies should generate novel insights into the molecular mechanism governing endosome-to-lysosome trafficking in neurons, and advance our understanding of the pathogenic mechanism of abnormal endosomal-lysosomal pathway in a variety of neurological disorders and neurodegenerative diseases.